Aseptic Technique: Media and Equipment

Growth Medium A growth medium or culture medium is a liquid or gel designed to _______ ____ _______ of microorganisms or cells The most common growth media for microorganisms are ________ _______ and _______ plates

Defined Media An important distinction between growth media types is that of ________ versus __________ media A defined medium will have _________ quantities of all ingredients

Undefined Media An undefined medium has some complex ingredients that consist of a mixture of many chemical species in _________ _____________ Undefined media are sometimes chosen based on _______ and sometimes by ____________ - some microorganisms have never been cultured on defined media.

Undefined Media We will use this in the lab Nutrient Agar contains: ________ _______ ___ ________ ________ ________ ________

Forms of Culture The most common growth media for microorganisms are ________ _______(liquid in a test tube) Liquid media are often mixed with ______ and poured into ______ _______ to solidify These agar plates provide a _______ medium on which microbes may be cultured. They remain solid, as very few bacteria are able to decompose agar.

Slants There are several reasons a slant is best for culture storage: 1) ________ ______ – slant increases it 2) ______ – test tubes smaller to store 3) ________ – won’t dry out 4) _______ – once the microorganism is transferred, you can store it for up to 6 months in a refrigerator

Aseptic Technique Aseptic technique refers to a procedure that is performed under ________ ____________ This includes ________ and _________ techniques (such as with microbiological cultures) It includes techniques like _________ ______________ (Bunsen burner)

Aseptic Technique Sterile surfaces must be protected from ________ ___ ____ ____ or on non-sterile surfaces In sterile technique, only ________ surfaces touch other ________ surfaces and ____ exposure is kept to a minimum

Aseptic Technique It is important in microbiology to work with ________ __________ This is difficult because the world around us is covered with ________ (even on dust particles in the air) In order to protect broth, plates, slants and pure cultures from MO’s around us, we practice _________ __________

Aseptic Technique In this class you will need to practice sterile technique when we _________ a pure culture into ______ __________ (tube of sterile broth or an agar plate)

Aseptic Technique “A” = Negative prefix “Septic” = Infection All techniques and procedures which __________ _______________

Safe Patterns Safe patterns include: Standard _______ _________ ______________ Techniques ______ and ________ of Equipment _________ Hands

Safe Patterns ________ all materials before beginning

Safe Patterns Spray the lab top down with a _________ _________ or a ______ _______ __________ and allow this to stand for a _______. You may then wipe down the bench with the paper towel.

Safe Patterns Wash hands _______ and _______ lab.

Safe Patterns You should have only the _________ ______ ________ and the written _____ __________ on your bench top or desk.

Safe Patterns Keep petri dishes and test tubes ___________ as much as possible. If top must be removed completely do ______ _____ _____on the lab top. This lowers the probability of contamination and prevents “false positive” results.

Safe Patterns Hold bottles and tubes ____ ___ _______ to minimize the amount of airborne microbes that can fall into them (blue circle). Remove the caps as shown above and ___ _____set the caps down. Keep the mouth of the cap facing _________ (red circle).

Safe Patterns When using metal ________ ________, HEAT the _______ piece of metal of the inoculation instrument in the flame: it should be RED HOT. Be sure to ________ your inoculation instrument ________ picking the inoculum (broth or agar).

Safe Patterns To __________ a Petri plate: Lift one edge of the Petri plate cover to gain access to the culture medium. Keep the cover over the plate bottom to prevent ______ and __________ from falling onto the agar.

Safe Patterns Report _________ to me immediately Cover the spill with ______ ________ and squirt _________ onto the towels. Wait _____ _________ then clean up the spill.

Safe Patterns ________ all test tubes and petri plates with your name (initials), date, and name of organism ________ you add any solutions, bacteria, etc. 

Safe Patterns Do _______ dump ANY microbial suspension down the ______ or in the _______ ______. I will collect them for proper disposal.

Safe Patterns Place _____ ________in racks when working at your table: never lay the tubes down—____ _____ Keep test tube ______ and petri dish _______ on media to reduce _______________ (matters not whether it is sterile media or already cultured).

Safe Patterns All agar plates are incubated _______ ______ to reduce bacterial _______________ and to reduce the possibility of water _______________ that may be on the lid dropping onto the agar, causing fluid to run across the agar medium.  

Media __________ after preparation, in storage, or working containers. Handle _____________. If no goofs – Sterile ___________

E. Autoclave _____________ ________ Water boils at 100ºC, but if pressure is __________ the boiling point _______ At 15 PSI the Boiling Point = 121ºC At 20 PSI the Boiling Point = 126ºC Minimum = ____ minutes @ ____ PSI A pressure cooker is the same as an ____________

Boiling _________ kills _______ MO, fungi, and viruses but a few are ____________. Example: Inf. Hepatitis, Endospores But very good nonprofessional use

Technique depends on article: Equipment Technique depends on article: Does it _____ or _____? Do you want it back?

Equipment Autoclave If it _____ into machine and ____ ____ ___ minutes @ 121ºC (15 PSI) minimum

Equipment Boiling ________ in a lab

Equipment Oven _____ ______ used for metal and glass ____ minutes @ _________ minimum

Equipment ___________ – Burning ________ Plus Loops, test tubes

Equipment _____________ Equipment Usually _______ and ______ materials Gas: _______ _______ ____________